Suppression of anoikis in human intestinal epithelial cells: differentiation state-selective roles of α2β1, α3β1, α5β1, and α6β4 integrins

BMC Cell Biol. 2013 Dec 1:14:53. doi: 10.1186/1471-2121-14-53.

Abstract

Background: Regulation of anoikis in human intestinal epithelial cells (IECs) implicates differentiation state-specific mechanisms. Human IECs express distinct repertoires of integrins according to their state of differentiation. Therefore, we investigated whether α2β1, α3β1, α5β1, and α6β4 integrins perform differentiation state-specific roles in the suppression of IEC anoikis.

Results: Human (HIEC, Caco-2/15) IECs were exposed to specific antibodies that block the binding activity of integrin subunits (α2, α3, α5, α6, β1 or β4) to verify whether or not their inhibition induced anoikis. The knockdown of α6 was also performed by shRNA. Additionally, apoptosis/anoikis was induced by pharmacological inhibition of Fak (PF573228) or Src (PP2). Anoikis/apoptosis was assayed by DNA laddering, ISEL, and/or caspase activity (CASP-8, -9, or -3). Activation levels of Fak and Src, as well as functional Fak-Src interactions, were also assessed. We report herein that differentiated IECs exhibit a greater sensitivity to anoikis than undifferentiated ones. This involves an earlier onset of anoikis when kept in suspension, as well as significantly greater contributions from β1 and β4 integrins in the suppression of anoikis in differentiated cells, and functional distinctions between β1 and β4 integrins in engaging both Fak and Src, or Src only, respectively. Likewise, Fak performs significantly greater contributions in the suppression of anoikis in differentiated cells. Additionally, we show that α2β1 and α5β1 suppress anoikis in undifferentiated cells, whereas α3β1 does so in differentiated ones. Furthermore, we provide evidence that α6β4 contributes to the suppression of anoikis in a primarily α6 subunit-dependent manner in undifferentiated cells, whereas this same integrin in differentiated cells performs significantly greater contributions in anoikis suppression than its undifferentiated state-counterpart, in addition to doing so through a dependence on both of its subunits.

Conclusions: Our findings indicate that the suppression of human IEC anoikis implicates differentiation state-selective repertoires of integrins, which in turn results into distinctions in anoikis regulation, and sensitivity, between undifferentiated and differentiated IECs. These data further the functional understanding of the concept that the suppression of anoikis is subjected to cell differentiation state-selective mechanisms.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anoikis / genetics*
  • Antibodies / pharmacology
  • Caco-2 Cells
  • Cell Differentiation
  • Cell Proliferation
  • Focal Adhesion Kinase 1 / antagonists & inhibitors
  • Focal Adhesion Kinase 1 / genetics
  • Focal Adhesion Kinase 1 / metabolism
  • Gene Expression Regulation
  • Humans
  • Integrin alpha2beta1 / antagonists & inhibitors
  • Integrin alpha2beta1 / genetics*
  • Integrin alpha2beta1 / metabolism
  • Integrin alpha3beta1 / antagonists & inhibitors
  • Integrin alpha3beta1 / genetics*
  • Integrin alpha3beta1 / metabolism
  • Integrin alpha5beta1 / antagonists & inhibitors
  • Integrin alpha5beta1 / genetics*
  • Integrin alpha5beta1 / metabolism
  • Integrin alpha6beta4 / antagonists & inhibitors
  • Integrin alpha6beta4 / genetics*
  • Integrin alpha6beta4 / metabolism
  • Intestinal Mucosa / metabolism*
  • Intestinal Mucosa / pathology
  • Protein Kinase Inhibitors
  • Pyrimidines / pharmacology
  • Quinolones / pharmacology
  • RNA, Small Interfering / genetics
  • RNA, Small Interfering / metabolism
  • Signal Transduction
  • Sulfones / pharmacology
  • src-Family Kinases / antagonists & inhibitors
  • src-Family Kinases / genetics
  • src-Family Kinases / metabolism

Substances

  • 6-(4-(3-(methylsulfonyl)benzylamino)-5-(trifluoromethyl)pyrimidin-2-ylamino)-3,4-dihydroquinolin-2(1H)-one
  • AG 1879
  • Antibodies
  • Integrin alpha2beta1
  • Integrin alpha3beta1
  • Integrin alpha5beta1
  • Integrin alpha6beta4
  • Protein Kinase Inhibitors
  • Pyrimidines
  • Quinolones
  • RNA, Small Interfering
  • Sulfones
  • Focal Adhesion Kinase 1
  • PTK2 protein, human
  • src-Family Kinases